Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, 8-1 Kawada-cho, Shinjuku-ku, Tokyo 162-8666, Japan.
World J Gastroenterol. 2009 Nov 14;15(42):5307-15. doi: 10.3748/wjg.15.5307.
To investigate the molecular mechanisms involved in coagulation factor expression and/or function during direct hyperplasia (DH)-mediated liver regeneration.
Direct hyperplasia-mediated liver regeneration was induced in female C57BL/6 mice by administering 1,4-bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP), a representative hepatomitogen. Mice were weighed and sacrificed at various time points [Day 0 (D0: prior to injection), 3 h, D1, D2, D3, and D10] after TCPOBOP administration to obtain liver and blood samples. Using the RNA samples extracted from the liver, a comprehensive analysis was performed on the hepatic gene expression profiling of coagulation-related factors by real-time RT-PCR (fibrinogen, prothrombin, factors V, VII, VIII, IX, X, XI, XII, XIIIbeta, plasminogen, antithrombin, protein C, protein S, ADAMTS13, and VWF). The corresponding plasma levels of coagulation factors (fibrinogen, prothrombin, factors V, VII, VIII, IX, X, XI, XII, XIII, and VWF) were also analyzed and compared with their mRNA levels.
Gavage administration of TCPOBOP (3 mg/kg body weight) resulted in a marked and gradual increase in the weight of the mouse livers relative to the total body weight to 220% by D10 relative to the D0 (control) ratios. At the peak of liver regeneration (D1 and D2), the gene expression levels for most of the coagulation-related factors (fibrinogen, prothrombin, factors V, VII, VIII, IX, XI, XII, XIIIbeta, plasminogen, antithrombin, protein C, ADAMTS13, VWF) were found to be down-regulated in a time-dependent manner, and gradually recovered by D10 to the basal levels. Only mRNA levels of factor X and protein S failed to show any decrease during the regenerative phase. As for the plasma levels, 5 clotting factors (prothrombin, factors VIII, IX, XI, and XII) demonstrated a significant decrease (P<0.05) during the regeneration phase compared with D0. Among these 5 factors, factor IX and factor XI showed the most dramatic decline in their activities by about 50% at D2 compared to the basal levels, and these reductions in plasma activity for both factors were consistent with our RT-PCR findings. In contrast, the plasma activities of the other coagulation factors (fibrinogen, factors V, VII, XIII, and VWF) were not significantly reduced, despite the reduction in the liver mRNA levels. Unlike the other factors, FX showed a temporal increase in its plasma activity, with significant increases (P<0.05) detected at D1.
Investigating the coagulation cascade protein profiles during liver regeneration by DH may help to better understand the basic biology of the liver under normal and pathological conditions.
研究直接增生（DH）介导的肝再生过程中凝血因子表达和/或功能涉及的分子机制。
通过给予 1,4-双[2-(3,5-二氯吡啶氧基)]苯（TCPOBOP），一种代表性的肝有丝分裂原，在雌性 C57BL/6 小鼠中诱导直接增生介导的肝再生。在 TCPOBOP 给药后，于不同时间点（D0：注射前、3 h、D1、D2、D3 和 D10）测量小鼠体重并处死，以获取肝脏和血液样本。使用从肝脏提取的 RNA 样本，通过实时 RT-PCR（纤维蛋白原、凝血酶原、因子 V、VII、VIII、IX、X、XI、XII、XIIIβ、纤溶酶原、抗凝血酶、蛋白 C、蛋白 S、ADAMTS13 和 VWF）对凝血相关因子的肝基因表达谱进行全面分析。还分析并比较了相应的血浆凝血因子（纤维蛋白原、凝血酶原、因子 V、VII、VIII、IX、X、XI、XII、XIII 和 VWF）水平与它们的 mRNA 水平。
TCPOBOP（3mg/kg 体重）灌胃给药后，与 D0（对照）相比，到 D10 时，小鼠肝脏相对于体重的重量比增加到 220%。在肝再生的高峰期（D1 和 D2），大多数凝血相关因子（纤维蛋白原、凝血酶原、因子 V、VII、VIII、IX、XI、XII、XIIIβ、纤溶酶原、抗凝血酶、蛋白 C、ADAMTS13、VWF）的基因表达水平呈时间依赖性下调，并在 D10 时逐渐恢复到基础水平。只有因子 X 和蛋白 S 的 mRNA 水平在再生阶段没有显示出任何下降。对于血浆水平，与 D0 相比，在再生阶段有 5 种凝血因子（凝血酶原、因子 VIII、IX、XI 和 XII）显著下降（P<0.05）。在这 5 种因子中，因子 IX 和因子 XI 的活性在 D2 时下降最明显，约为基础水平的 50%，这两种因子的血浆活性降低与我们的 RT-PCR 结果一致。相比之下，其他凝血因子（纤维蛋白原、因子 V、VII、XIII 和 VWF）的血浆活性没有明显降低，尽管肝脏的 mRNA 水平降低。与其他因子不同，FX 的血浆活性呈时间性增加，在 D1 时检测到显著增加（P<0.05）。
通过 DH 研究肝再生过程中的凝血级联蛋白谱可能有助于更好地理解正常和病理条件下肝脏的基本生物学。
